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Introduction: Unlike idiopathic nephrotic syndrome (NS), hereditary podocytopathies are not expected to recur after kidney transplantation. However, some reports of posttransplant recurrence of NS in patients carrying variants in the NPHS2 gene have been described, notably with the p.Arg138Gln variant, which is more prevalent in Europe. The objective of this study was to assess the risk of recurrence after kidney transplantation in a large cohort of patients with biallelic NPHS2 pathogenic variants. Methods: Since January 2010, 61 patients identified at Necker-Enfants Malades Hospital and 56 enrolled in the PodoNet Registry with biallelic variants in the NPHS2 gene were transplanted and were compared with 44 transplanted children with steroid-resistant NS (SRNS) without any identified pathogenic variant. Results: Of the 117 patients, 23 carried the p.Arg138Gln variant in the homozygous state and 16 in the compound heterozygous state. The other 78 patients carried different variants in the homozygous (n = 44) or compound heterozygous state. Only 1 patient with NPHS2-related SRNS experienced posttransplant recurrence (median follow-up of cohort 8.5 years [2.5-15]). Conversely, 7 of 44 patients (16%) without any identified pathogenic variant recurred within a maximum of 7 days after transplantation (median follow-up 8.9 years [0.6-13.9]). Conclusion: In this large cohort, the risk of patients with causative variants in the NPHS2 gene to develop NS recurrence after kidney transplantation was extremely low. This is coherent with the pathophysiology of intrinsic slit-diaphragm disease. These data are reassuring and should be considered when counselling patients, making living kidney donation, whether related or not, a safe choice.
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The association of both uniparental disomy and small supernumerary marker chromosomes is rare. Clinical impact depends on the presence of imprinted genes and/or the unmasking of a recessive mutation of the chromosome involved in the uniparental disomy and the euchromatic content of the sSMC. Here, we report on the second case of a patient harbouring a de novo supernumerary marker chromosome 6 causing partial trisomy 6p12.3p11.1 associated with a paternal uniparental isodisomy of chromosome 6. Our patient presented with intrauterine growth retardation, macroglossia, initial developmental delay and transient neonatal diabetes mellitus followed by a congenital hyperinsulinism. Diabetes and intrauterine growth retardation can be linked to the paternal isodisomy of the imprinted locus on chromosome 6q24 whereas developmental delay is probably due to the small supernumerary marker chromosome. However, the clinical impact of partial trisomy 6p is difficult to address due to a limited number of patients. The careful clinical examination and the molecular characterization of additional patients with trisomy 6p are needed to further predict the phenotype for genetic counselling. Finally, uniparental disomy should be considered when a sSMC involving a chromosome containing imprinted regions is detected, especially in the prenatal setting.
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The human genome comprises approximately 3% of tandem repeats with variable length (VNTR), a few of which have been linked to human rare diseases. Autosomal dominant tubulointerstitial kidney disease-MUC1 (ADTKD-MUC1) is caused by specific frameshift variants in the coding VNTR of the MUC1 gene. Calling variants from VNTR using short-read sequencing (SRS) is challenging due to poor read mappability. We developed a computational pipeline, VNtyper, for reliable detection of MUC1 VNTR pathogenic variants and demonstrated its clinical utility in two distinct cohorts: (1) a historical cohort including 108 families with ADTKD and (2) a replication naive cohort comprising 2,910 patients previously tested on a panel of genes involved in monogenic renal diseases. In the historical cohort all cases known to carry pathogenic MUC1 variants were re-identified, and a new 25bp-frameshift insertion in an additional mislaid family was detected. In the replication cohort, we discovered and validated 30 new patients.
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X-linked Alport syndrome (XLAS) is an inherited kidney disease caused exclusively by pathogenic variants in the COL4A5 gene. In 10-20% of cases, DNA sequencing of COL4A5 exons or flanking regions cannot identify molecular causes. Here, our objective was to use a transcriptomic approach to identify causative events in a group of 19 patients with XLAS without identified mutation by Alport gene panel sequencing. Bulk RNAseq and/or targeted RNAseq using a capture panel of kidney genes was performed. Alternative splicing events were compared to those of 15 controls by a developed bioinformatic score. When using targeted RNAseq, COL4A5 coverage was found to be 23-fold higher than with bulk RNASeq and revealed 30 significant alternative splicing events in 17 of the 19 patients. After computational scoring, a pathogenic transcript was found in all patients. A causative variant affecting COL4A5 splicing and absent in the general population was identified in all cases. Altogether, we developed a simple and robust method for identification of aberrant transcripts due to pathogenic deep-intronic COL4A5 variants. Thus, these variants, potentially targetable by specific antisense oligonucleotide therapies, were found in a high percentage of patients with XLAS in whom pathogenic variants were missed by conventional DNA sequencing.
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Nefritis Hereditaria , Humanos , Nefritis Hereditaria/diagnóstico , Nefritis Hereditaria/genética , Nefritis Hereditaria/patología , Colágeno Tipo IV/genética , Colágeno Tipo IV/metabolismo , Mutación , Exones , Empalme del ARNRESUMEN
CHARGE syndrome, due to CHD7 pathogenic variations, is an autosomal dominant disorder characterized by a large spectrum of severity. Despite the great number of variations reported, no clear genotype-to-phenotype correlation has been reported. Unsupervised machine learning and clustering was undertaken using a retrospective cohort of 42 patients, after deep radiologic and clinical phenotyping, to establish genotype-phenotype correlation for CHD7-related CHARGE syndrome. It resulted in three clusters showing phenotypes of different severities. While no clear genotype-phenotype correlation appeared within the first two clusters, a single patient was outlying the cohort data (cluster 3) with the most atypical phenotype and the most distal frameshift variant in the gene. We added two other patients with similar distal pathogenic variants and observed a tendency toward mild and/or atypical phenotypes. We hypothesized that this finding could potentially be related to escaping nonsense mediated RNA decay, but found no evidence of such decay in vivo for any of the CHD7 pathogenic variation tested. This indicates that this milder phenotype may rather result from the production of a protein retaining all functional domains.
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Síndrome CHARGE , Humanos , Síndrome CHARGE/genética , Estudios Retrospectivos , Fenotipo , Estudios de Asociación Genética , Genotipo , Mutación/genéticaRESUMEN
The rise of genetics in the last decades has allowed major advances in the understanding of the mechanisms leading to inherited kidney diseases. From the first positional cloning studies to the advent of high-throughput sequencing (NGS), genome analysis technologies have become increasingly efficient, with an extraordinary level of resolution. Moreover, sequencing prices have decreased from one million dollars for the sequencing of James Watson's genome in 2008, to a few hundred dollars for the sequencing of a genome today. Thus, molecular diagnosis has a central place in the diagnosis of these patients and influences the therapeutic management in many situations. However, although NGS is a powerful tool for the identification of variants involved in diseases, it also exposes to the risk of over-interpretation of certain variants, leading to erroneous diagnoses, requiring the use of specialists. In this review, we first propose a brief retrospective of the essential steps that led to the current knowledge and the development of NGS for the study of hereditary nephropathies in children. This review is then an opportunity to present the main hereditary nephropathies and the underlying molecular mechanisms. Among them, we emphasize ciliopathies, congenital anomalies of the kidney and urinary tract, podocytopathies and tubulopathies.
Title: Les grandes avancées en néphro-génétique pédiatrique. Abstract: L'essor de la génétique au cours des dernières décennies a permis des avancées majeures dans la compréhension des mécanismes conduisant aux maladies rénales héréditaires. Des premières études par clonage positionnel jusqu'à l'avènement du séquençage à haut débit (NGS), les techniques d'analyse du génome sont devenues de plus en plus performantes, avec un niveau de résolution extraordinaire. Les prix de séquençage se sont effondrés, passant d'un million de dollars (environ 940 millions d'euros) pour le séquençage du génome de James Watson en 2008, à quelques centaines d'euros pour le séquençage d'un génome aujourd'hui. Le diagnostic moléculaire tient ainsi une place centrale pour le diagnostic des patients et influe sur la prise en charge thérapeutique dans de nombreuses situations. Mais si le NGS est un outil performant pour l'identification de variants impliqués dans les maladies, il expose au risque de surinterprétation de certains variants, conduisant à des diagnostics erronés. Dans cette revue, nous proposons une brève rétrospective des étapes essentielles qui ont conduit aux connaissances actuelles et au développement du NGS pour l'étude des néphropathies héréditaires de l'enfant. Nous développerons ensuite les principales néphropathies héréditaires et les mécanismes moléculaires sous-jacents.
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Ciliopatías , Secuenciación de Nucleótidos de Alto Rendimiento , Niño , Humanos , Estudios RetrospectivosRESUMEN
Genetic studies of hereditary steroid resistant nephrotic syndrome (SRNS) have identified more than 60 genes involved in the development of single-gene, isolated or syndromic forms of hereditary podocytoapthies. Sometimes, syndromic SRNS is associated with neurological disorders. Over the past decades, various studies have established links between the podocyte, an epithelial glomerular cell involved in the renal filtration barrier, and neuronal cells, both morphologically (slit diaphragm and synapse) and functionally (signaling platforms). Variants of genes encoding proteins expressed in different compartments of the podocyte and neurons are responsible for phenotypes associating renal lesions with proteinuria to central and/or peripheral neurological disorders. In this review, we aim to focus on genetic syndromes associating proteinuria and neurological disease and to present the latest advances in the description of these neuro-renal disorders.
Title: Atteinte neurologique et syndrome néphrotique cortico-résistant - Des chevauchements troublants. Abstract: Les études génétiques portant sur le syndrome néphrotique (SN) héréditaire ont permis d'identifier plus de 60 gènes impliqués dans le développement de formes monogéniques de SN cortico-résistant, isolé ou syndromique, ce dernier étant parfois associé à des troubles neurologiques. Au cours des dernières décennies, diverses études ont établi des liens entre la physiologie des podocytes et celle des neurones, tant sur le plan morphologique (diaphragme de fente et synapse) que fonctionnel (plateformes de signalisation). Des variants dans des gènes codant des protéines s'exprimant dans différents compartiments du podocyte et des neurones sont responsables de phénotypes associant des lésions rénales avec protéinurie à des troubles neurologiques centraux et/ou périphériques. L'objectif de cette revue est de se concentrer sur les syndromes génétiques associant une protéinurie et une atteinte neurologique et de présenter les dernières avancées dans la description de ces troubles neuro-rénaux.
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Síndrome Nefrótico , Enfermedades del Sistema Nervioso , Humanos , Síndrome Nefrótico/genética , Glomérulos Renales/patología , Riñón/patología , Proteinuria , Enfermedades del Sistema Nervioso/genéticaRESUMEN
Whole-genome sequencing (WGS) now allows identification of multiple variants in non-coding regions. The large number of variants identified by WGS however complicates their interpretation. Through identification of the first deep intronic variant in NPHS2, which encodes podocin, a protein implicated in autosomal recessive steroid resistant nephrotic syndrome (SRNS), we compare herein three different tools including a newly developed targeted NGS-based RNA-sequencing to explore the splicing effect of intronic variations. WGS identified two different variants in NPHS2 eventually involved in the disease. Through RT-PCR, exon-trapping Minigene assay and targeted RNA sequencing, we were able to identify the splicing defect in NPHS2 mRNA from patient kidney tissue. Only targeted RNA-seq simultaneously analyzed the effect of multiple variants and offered the opportunity to quantify consequences on splicing. Identifying deep intronic variants and their role in disease is of utmost importance. Alternative splicing can be predicted by in silico tools but always requires confirmation through functional testing with RNA analysis from the implicated tissue remaining the gold standard. When several variants with potential effects on splicing are identified by WGS, a targeted RNA sequencing panel could be of great value.
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Síndrome Nefrótico , Humanos , Mutación , Secuenciación Completa del Genoma , Síndrome Nefrótico/genética , ARN Mensajero/genéticaRESUMEN
BACKGROUND: Long-term outcomes after multiple courses of rituximab among children with frequently relapsing, steroid-dependent nephrotic syndrome (FRSDNS) are unknown. METHODS: A retrospective cohort study at 16 pediatric nephrology centers from ten countries in Asia, Europe, and North America included children with FRSDNS who received two or more courses of rituximab. Primary outcomes were relapse-free survival and adverse events. RESULTS: A total of 346 children (age, 9.8 years; IQR, 6.6-13.5 years; 73% boys) received 1149 courses of rituximab. A total of 145, 83, 50, 28, 22, and 18 children received two, three, four, five, six, and seven or more courses, respectively. Median (IQR) follow-up was 5.9 (4.3-7.7) years. Relapse-free survival differed by treatment courses (clustered log-rank test P<0.001). Compared with the first course (10.0 months; 95% CI, 9.0 to 10.7 months), relapse-free period and relapse risk progressively improved after subsequent courses (12.0-16.0 months; HRadj, 0.03-0.13; 95% CI, 0.01 to 0.18; P<0.001). The duration of B-cell depletion remained similar with repeated treatments (6.1 months; 95% CI, 6.0 to 6.3 months). Adverse events were mostly mild; the most common adverse events were hypogammaglobulinemia (50.9%), infection (4.5%), and neutropenia (3.7%). Side effects did not increase with more treatment courses nor a higher cumulative dose. Only 78 of the 353 episodes of hypogammaglobulinemia were clinically significant. Younger age at presentation (2.8 versus 3.3 years; P=0.05), age at first rituximab treatment (8.0 versus 10.0 years; P=0.01), and history of steroid resistance (28% versus 18%; P=0.01) were associated with significant hypogammaglobulinemia. All 53 infective episodes resolved, except for one patient with hepatitis B infection and another with EBV infection. There were 42 episodes of neutropenia, associated with history of steroid resistance (30% versus 20%; P=0.04). Upon last follow-up, 332 children (96%) had normal kidney function. CONCLUSIONS: Children receiving repeated courses of rituximab for FRSDNS experience an improving clinical response. Side effects appear acceptable, but significant complications can occur. These findings support repeated rituximab use in FRSDNS.
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Agammaglobulinemia , Nefrosis Lipoidea , Síndrome Nefrótico , Neutropenia , Agammaglobulinemia/inducido químicamente , Agammaglobulinemia/tratamiento farmacológico , Niño , Femenino , Humanos , Inmunosupresores/uso terapéutico , Masculino , Nefrosis Lipoidea/tratamiento farmacológico , Síndrome Nefrótico/tratamiento farmacológico , Neutropenia/inducido químicamente , Neutropenia/tratamiento farmacológico , Recurrencia , Estudios Retrospectivos , Rituximab/efectos adversos , Esteroides/uso terapéutico , Resultado del TratamientoRESUMEN
We report the screening of a large panel of genes in a series of 100 fetuses (98 families) affected with severe renal defects. Causative variants were identified in 22% of cases, greatly improving genetic counseling. The percentage of variants explaining the phenotype was different according to the type of phenotype. The highest diagnostic yield was found in cases affected with the ciliopathy-like phenotype (11/15 families and, in addition, a single heterozygous or a homozygous Class 3 variant in PKHD1 in three unrelated cases with autosomal recessive polycystic kidney disease). The lowest diagnostic yield was observed in cases with congenital anomalies of the kidney and urinary tract (9/78 families and, in addition, Class 3 variants in GREB1L in three unrelated cases with bilateral renal agenesis). Inheritance was autosomal recessive in nine genes (PKHD1, NPHP3, CEP290, TMEM67, DNAJB11, FRAS1, ACE, AGT, and AGTR1), and autosomal dominant in six genes (PKD1, PKD2, PAX2, EYA1, BICC1, and MYOCD). Finally, we developed an original approach of next-generation sequencing targeted RNA sequencing using the custom capture panel used for the sequencing of DNA, to validate one MYOCD heterozygous splicing variant identified in two male siblings with megabladder and inherited from their healthy mother.
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Enfermedades Renales , Riñón Poliquístico Autosómico Dominante , Antígenos de Neoplasias , Proteínas de Ciclo Celular/genética , Proteínas del Citoesqueleto/genética , Femenino , Feto/anomalías , Secuenciación de Nucleótidos de Alto Rendimiento , Homocigoto , Humanos , Riñón/anomalías , Enfermedades Renales/congénito , Enfermedades Renales/diagnóstico , Enfermedades Renales/genética , Masculino , Mutación , Riñón Poliquístico Autosómico Dominante/genéticaRESUMEN
BACKGROUND: A precise assessment of glomerular filtration rate is key to delineate the care of children with a solitary functioning kidney (SFK). Data regarding measured GFR (mGFR) in this population is restricted to a single study of 77 individuals, which suggested that a GFR estimation (eGFR) method based on creatinine and cystatin C (eGFR-CKiD2) performed better than Schwartz's equation (eGFR-Schwartz). METHODS: We measured GFR in 210 consecutive adolescents (7 to 22 years old) with an SFK referred to our institution between 2014 and 2019 and in 43 young candidates for kidney donation (18 to 25 years old). We compared the distribution of mGFR in both groups and determined the factors associated with reduced mGFR in adolescents with an SFK. We further compared different eGFR formulas with mGFR and assessed the association of mGFR and eGFRs with PTH and FGF23, two early indicators of GFR reduction. RESULTS: While adolescents with an SFK had a similar median mGFR to healthy controls (103 ± 24ml/min/1.73m2 vs. 107 ± 12 ml/min/1.73m2), the fraction of individuals with an mGFR below 90 ml/min/1.73m2 was higher in patients with SFK (23% vs. 5% in controls; P = 0.005). Multiple linear regression identified older age, ipsilateral abnormalities of the urinary tract, lack of compensatory hypertrophy, and treated hypertension as independent factors associated with reduced mGFR. A smaller bias using eGFR-Schwartz (95% confidence interval (95%CI): 3 to 7) was revealed when compared to other eGFR. Compared to eGFR-Schwartz, mGFR showed a stronger correlation with PTH (r = 0.04 vs. r = 0.1) and FGF23 (r = 0.03 vs. r = 0.05). CONCLUSION: SFK is not a benign condition, since 20% of the patients display altered kidney function. Our results raise caution regarding the use of the cystatin-based equation. mGFR shows a better ability than eGFR-Schwartz to differentiate patients showing early homeostatic adaptation to GFR reduction.